Myosin light-chain kinase can be phosphorylated by the catalytic subunit of cyclic AMP-dependent protein kinase. Phosphorylation of myosin kinase decreases the affinity of this enzyme for the calcium binding protein calmodulin by twenty fold. The ability of cyclic AMP dependent protein kinase to decrease the activity of myosin light chain kinase, provides a direct mechanism for cyclic AMP mediated smooth muscle relaxation.